Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping
Abstract Background The detection and identification of single nucleotide polymorphism (SNP) is essential for determining patient disease susceptibility and the delivery of medicines targeted to the individual. At present, SNP genotyping technology includes Sanger sequencing, TaqMan-probe quantitati...
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| Format: | Article |
| Language: | English |
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BMC
2021-11-01
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| Series: | BMC Genomics |
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| Online Access: | https://doi.org/10.1186/s12864-021-08148-2 |
| _version_ | 1819126979512762368 |
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| author | Baowei Li Yanran Liu Xiaodan Hao Jinhua Dong Limei Chen Haimei Li Wei Wu Ying Liu Jianxun Wang Yin Wang Peifeng Li |
| author_facet | Baowei Li Yanran Liu Xiaodan Hao Jinhua Dong Limei Chen Haimei Li Wei Wu Ying Liu Jianxun Wang Yin Wang Peifeng Li |
| author_sort | Baowei Li |
| collection | DOAJ |
| description | Abstract Background The detection and identification of single nucleotide polymorphism (SNP) is essential for determining patient disease susceptibility and the delivery of medicines targeted to the individual. At present, SNP genotyping technology includes Sanger sequencing, TaqMan-probe quantitative polymerase chain reaction (qPCR), amplification-refractory mutation system (ARMS)-PCR, and Kompetitive Allele-Specific PCR (KASP). However, these technologies have some disadvantages: the high cost of development and detection, long and time consuming protocols, and high false positive rates. Focusing on these limitations, we proposed a new SNP detection method named universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR). In this method, only two types of fluorescence-labeled probes were used for SNP genotyping, thus greatly reducing the cost of development and detection for SNP genotyping. Results In the amplification process of UPIP-qPCR, unlabeled intermediate primers with template-specific recognition functions could trigger probe hydrolysis and specific signal release. UPIP-qPCR can be used successfully and widely for SNP genotyping. The sensitivity of UPIP-qPCR in SNP genotyping was 0.01 ng, the call rate was more than 99.1%, and the accuracy was more than 99.9%. High-throughput DNA microarrays based on intermediate primers can be used for SNP genotyping. Conclusion This novel approach is both cost effective and highly accurate; it is a reliable SNP genotyping method that would serve the needs of the clinician in the provision of targeted medicine. |
| first_indexed | 2024-12-22T08:04:38Z |
| format | Article |
| id | doaj.art-6e04b79e072348569f454e7bbdd7f48c |
| institution | Directory Open Access Journal |
| issn | 1471-2164 |
| language | English |
| last_indexed | 2024-12-22T08:04:38Z |
| publishDate | 2021-11-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Genomics |
| spelling | doaj.art-6e04b79e072348569f454e7bbdd7f48c2022-12-21T18:33:10ZengBMCBMC Genomics1471-21642021-11-0122111110.1186/s12864-021-08148-2Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotypingBaowei Li0Yanran Liu1Xiaodan Hao2Jinhua Dong3Limei Chen4Haimei Li5Wei Wu6Ying Liu7Jianxun Wang8Yin Wang9Peifeng Li10Key Laboratory of Biological Medicines in Universities of Shandong Province, Weifang Key Laboratory of Antibody Medicines, School of Bioscience and Technology, Weifang Medical UniversitySchool of Basic Medicine, Qingdao UniversityInstitute for Translational Medicine, Qingdao UniversityKey Laboratory of Biological Medicines in Universities of Shandong Province, Weifang Key Laboratory of Antibody Medicines, School of Bioscience and Technology, Weifang Medical UniversityKey Laboratory of Biological Medicines in Universities of Shandong Province, Weifang Key Laboratory of Antibody Medicines, School of Bioscience and Technology, Weifang Medical UniversityKey Laboratory of Biological Medicines in Universities of Shandong Province, Weifang Key Laboratory of Antibody Medicines, School of Bioscience and Technology, Weifang Medical UniversityCollege of Food Science and Engineering, Qingdao Agricultural UniversityInstitute for Translational Medicine, Qingdao UniversitySchool of Basic Medicine, Qingdao UniversityInstitute for Translational Medicine, Qingdao UniversityInstitute for Translational Medicine, Qingdao UniversityAbstract Background The detection and identification of single nucleotide polymorphism (SNP) is essential for determining patient disease susceptibility and the delivery of medicines targeted to the individual. At present, SNP genotyping technology includes Sanger sequencing, TaqMan-probe quantitative polymerase chain reaction (qPCR), amplification-refractory mutation system (ARMS)-PCR, and Kompetitive Allele-Specific PCR (KASP). However, these technologies have some disadvantages: the high cost of development and detection, long and time consuming protocols, and high false positive rates. Focusing on these limitations, we proposed a new SNP detection method named universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR). In this method, only two types of fluorescence-labeled probes were used for SNP genotyping, thus greatly reducing the cost of development and detection for SNP genotyping. Results In the amplification process of UPIP-qPCR, unlabeled intermediate primers with template-specific recognition functions could trigger probe hydrolysis and specific signal release. UPIP-qPCR can be used successfully and widely for SNP genotyping. The sensitivity of UPIP-qPCR in SNP genotyping was 0.01 ng, the call rate was more than 99.1%, and the accuracy was more than 99.9%. High-throughput DNA microarrays based on intermediate primers can be used for SNP genotyping. Conclusion This novel approach is both cost effective and highly accurate; it is a reliable SNP genotyping method that would serve the needs of the clinician in the provision of targeted medicine.https://doi.org/10.1186/s12864-021-08148-2UPIP-qPCRIntermediate primerUniversal probeSNP genotypingSanger sequencing |
| spellingShingle | Baowei Li Yanran Liu Xiaodan Hao Jinhua Dong Limei Chen Haimei Li Wei Wu Ying Liu Jianxun Wang Yin Wang Peifeng Li Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping BMC Genomics UPIP-qPCR Intermediate primer Universal probe SNP genotyping Sanger sequencing |
| title | Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping |
| title_full | Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping |
| title_fullStr | Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping |
| title_full_unstemmed | Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping |
| title_short | Universal probe-based intermediate primer-triggered qPCR (UPIP-qPCR) for SNP genotyping |
| title_sort | universal probe based intermediate primer triggered qpcr upip qpcr for snp genotyping |
| topic | UPIP-qPCR Intermediate primer Universal probe SNP genotyping Sanger sequencing |
| url | https://doi.org/10.1186/s12864-021-08148-2 |
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